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TRACK 3 TRACK 3 Technical Program

sity of regenerative medicine applications. Extracellular matrix (ECM) rep- verge to form local gaps which contrary to findings from 2D models, close
resents an essential player in stem cell niche, since it can directly or indirect- faster from regions of high curvature. Gap closure kinetics is regulated by
ly modulate the maintenance, proliferation, self-renewal and differentiation gap size and shape, with gaps <85,000 µm2 closing and >> 140,000 µm2
of stem cells. When creating 3D scaffolds for cells, it is preferable to provide not closing regardless of shape, while shape influencing closure of those in
cells with an environment which closely resembles their native ECM. between. The inhibition of actin-myosin interactions through treatment with
Electrospinning has been proved to be an effective technique for the con- 10 µM and 20 µM y-27632 compromised the closure of large local gaps
trolled fabrication of micro-fiber meshes for tissue engineering applications, (>>40,000µm2). However, small local gaps (<10,000µm2) closed despite
due to its ability to produce biocompatible and biomimetic scaffolds capable exposure to the drug. Overall gap closure was significantly influenced by
of supporting cell growth. fiber architecture with crosshatch patterned fibers closing the gaps faster
when compared to gaps bridged by parallel array of fibers. We have system-
Herein we proposed an electrospun gelatin based scaffold to resemble the atically studied the emergence of cells from monolayers initiating formation
ECM topography, for the sustainment of stem cells survival, proliferation and of cell streams, and SCS, thus facilitating the overall collective cell migration
differentiation. We characterized the scaffold using scanning electron mi- process. Given the observed influence of size and fiber architecture on gap
croscopy, Fourier transform infrared spectroscopy, and confocal microscopy. closure, findings from this study could be utilized to design in vitro scaffolds
We tested the biocompatibility of the electrospun scaffolds using human that serve as a foundation to generate non-closing diabetic wound, and or
MSC (hMSC). Early passage hMSC were seeded onto the scaffold to as- to study metastatic events in cancer biology.
sess cell growth up to one month. The results of the in vitro tests indicated
uncompromised cell viability and full cell adhesion to the patch for all the
duration of the experiment. Multilineage potential of cells toward osteogenic 3-8
and chondrogenic differentiation was assessed in vitro for up to 1 month.
The differentiation of hMSC was carried out using chondrogenic and osteo- TISSUE MODELS
genic inducing factors. SEM and confocal microscopy analysis were used to
compare the morphology of cells. Cytoskeleton morphology was assessed Bexar/Travis 4:00pm - 5:40pm
by fluorescent staining; cells on the patch exhibited spread morphology
with abundant cytoskeleton staining. Our data show that when appropriately
induced, MSC seeded onto the electrospun scaffold have the ability to dif- Session Organizer: Dan Dongeun Huh, Department of Bioengi-
ferentiate into the osteogenic and chondrogenic lineages; on the other side, neering, University of Pennsylvania, Philadelphia, PA, United States
hMSC harvested with the untreated culture media maintain their immuno-
suppressive potential and immunomodulatory properties. 4:00pm Microengineered cell and tissue systems: Evolution of
in-vitro liver technologies
A scaffold which mimics the native ECM while allowing for cell infiltration and
proliferation is important for regenerative medicine applications to permit
the penetration of host cells and vasculature, augmenting diffusion of nu- Technical Presentation. NEMB2016-5969
trients and waste products and enhancing integration into the host tissue.
Future application of this versatile scaffold platform to induced pluripotent Osman Berk USTA, Harvard Medical School - Massachusetts Gen-
stem cells for functional tissue repair and regeneration will further expand its eral Hospital, Boston, MA, United States
potential for regenerative therapies.
The liver performs many key functions such as serving as the metabolic hub
10:38am A Suspended and Aligned Nanofiber Network Assay of the body. For this reason, the liver is the focal point of many investigations
to Study Collective Cell Migration and Gap Closure Dynamics aimed at understanding an organism’s toxicological response to endoge-
nous and exogenous challenges. We will present a survey and critical com-
Technical Presentation. NEMB2016-6125 parison of in-vitro liver technologies along a broad spectrum, but focus on
the current renewed push to develop “organs-on-a-chip” in our laboratory
and elsewhere.
Puja Sharma, Virginia Tech, Blacksburg, VA, United States, Jerry
Lee, National Cancer Institute, NIH, Bethesda, MD, United States, 4:20pm Microfabricated Co-Cultures Containing Induced Plu-
Bahareh Behkam, Amrinder Nain, Virginia Tech, Blacksburg, VA, ripotent Stem Cell-Derived Liver Cells for Drug Development
United States
Technical Presentation. NEMB2016-6002
Collective cell migration described as coordinated movement of multiple
cells with well-maintained cell-cell adhesions is relevant to a myriad of
physiological phenomenon including wound healing, morphogenesis and Brenton Ware, Salman R. Khetani, University of Illinois at Chicago,
metastasis. In certain physiological conditions involving poorly developed Chicago, IL, United States
or absent extra-cellular matrix (ECM), collective cell migration is observed in
the form of suspended sheets. Our understanding of suspended cell migra- Introduction: Drug-induced liver injury is a leading cause of drug attrition in
tion is limited, as traditional wound healing scratch tests are conducted on the pharmaceutical industry. Significant differences in liver pathways across
featureless 2D substrates, which do not faithfully recapitulate the behavior species now necessitate the use of human-relevant cultures for assessing
of cells in their native fibrous ECM environment. Here, we present a novel liver-drug interactions. While primary human hepatocytes (PHHs) isolated
gap closure assay system comprised of suspended and aligned arrays (in from the liver are considered ideal for such purposes, these cells are in
parallel or crosshatch configurations) of ECM mimicking 500 nm diameter fi- limited supply for screening large (millions) libraries of drugs. Furthermore,
bers bridging the gap between two fibroblast monolayers cultured on raised lack of genetic diversity restricts the use of PHHs for identifying mecha-
platforms separated by a few millimeters. Fibroblasts sense fiber alignment nisms underlying inter-individual susceptibility to drugs. Induced pluripotent
and emerge from the monolayers as either individual leaders showing elas- stem cell-derived human hepatocyte-like cells (iHeps) have the potential to
tic recoils or as chains or as collective groups with intact cell-cell adhesions. address the aforementioned limitations with PHHs. However, iHeps remain
Using time-lapse microscopy, we find that the emerging cells form streams, fetal-like (i.e. low adult liver functions) under conventional culture formats
which advance rapidly (200µm/day) for a few days followed by slow growth that rely exclusively on extracellular matrices and soluble factors. Here, we
(20µm/day). Suspended cell sheets (SCS) formed between the cells streams, utilized soft lithography and co-culture with stromal cells to develop and
advance away from the monolayer in oscillatory patterns, with neighboring characterize a micropatterned co-culture (iMPCC) platform that can further
sheets demonstrating advancement in out-of-phase mode. SCS advance- mature iHeps towards an adult liver phenotype and maintain these functions 43
ment is inversely related to its span distance, with advancement severely for at least 4 weeks in vitro. We then tested toxicity of a panel of drugs in this
compromised at span distances exceeding 375µm. Advancing SCS con- platform and compared results to both PHH cultures and clinical findings.